RESUMO
Spotted fever group (SFG) rickettsiae are human pathogens that infect cells in the vasculature. They disseminate through host tissues by a process of cell-to-cell spread that involves protrusion formation, engulfment, and vacuolar escape. Other bacterial pathogens rely on actin-based motility to provide a physical force for spread. Here, we show that SFG species Rickettsia parkeri typically lack actin tails during spread and instead manipulate host intercellular tension and mechanotransduction to promote spread. Using transposon mutagenesis, we identified surface cell antigen 4 (Sca4) as a secreted effector of spread that specifically promotes protrusion engulfment. Sca4 interacts with the cell-adhesion protein vinculin and blocks association with vinculin's binding partner, α-catenin. Using traction and monolayer stress microscopy, we show that Sca4 reduces vinculin-dependent mechanotransduction at cell-cell junctions. Our results suggest that Sca4 relieves intercellular tension to promote protrusion engulfment, which represents a distinctive strategy for manipulating cytoskeletal force generation to enable spread.
Assuntos
Antígenos de Bactérias/metabolismo , Proteínas de Bactérias/metabolismo , Interações Hospedeiro-Patógeno , Mecanotransdução Celular , Infecções por Rickettsia/metabolismo , Infecções por Rickettsia/microbiologia , Rickettsia/patogenicidade , Vinculina/metabolismo , Actinas/metabolismo , Sequência de Aminoácidos , Antígenos de Bactérias/genética , Proteínas de Bactérias/genética , Caderinas/metabolismo , Adesão Celular , Linhagem Celular Tumoral , Elementos de DNA Transponíveis/genética , Febre/metabolismo , Febre/microbiologia , Humanos , Mutagênese Insercional , Mutação , Rickettsia/metabolismo , alfa Catenina/metabolismoRESUMO
Antigenically sequence variable M proteins of the major bacterial pathogen Streptococcus pyogenes (Strep A) are responsible for recruiting human C4b-binding protein (C4BP) to the bacterial surface, which enables Strep A to evade destruction by the immune system. The most sequence divergent portion of M proteins, the hypervariable region (HVR), is responsible for binding C4BP. Structural evidence points to the conservation of two C4BP-binding sequence patterns (M2 and M22) in the HVR of numerous M proteins, with this conservation applicable to vaccine immunogen design. These two patterns, however, only partially explain C4BP binding by Strep A. Here, we identified several M proteins that lack these patterns but still bind C4BP and determined the structures of two, M68 and M87 HVRs, in complex with a C4BP fragment. Mutagenesis of these M proteins led to the identification of amino acids that are crucial for C4BP binding, enabling formulation of new C4BP-binding patterns. Mutagenesis was also carried out on M2 and M22 proteins to refine or generate experimentally grounded C4BP-binding patterns. The M22 pattern was the most prevalent among M proteins, followed by the M87 and M2 patterns, while the M68 pattern was rare. These patterns, except for M68, were also evident in numerous M-like Enn proteins. Binding of C4BP via these patterns to Enn proteins was verified. We conclude that C4BP-binding patterns occur frequently in Strep A strains of differing M types, being present in their M or Enn proteins, or frequently both, providing further impetus for their use as vaccine immunogens.
Assuntos
Antígenos de Bactérias , Proteína de Ligação ao Complemento C4b , Streptococcus pyogenes , Streptococcus pyogenes/metabolismo , Streptococcus pyogenes/genética , Streptococcus pyogenes/química , Proteína de Ligação ao Complemento C4b/metabolismo , Antígenos de Bactérias/metabolismo , Antígenos de Bactérias/química , Antígenos de Bactérias/genética , Humanos , Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas da Membrana Bacteriana Externa/química , Proteínas da Membrana Bacteriana Externa/genética , Proteínas de Transporte/metabolismo , Proteínas de Transporte/genética , Proteínas de Transporte/química , Ligação Proteica , Sequência de Aminoácidos , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genéticaRESUMO
Enterococcus faecalis is an opportunistic pathogen frequently causing nosocomial infections. The virulence of this organism is underpinned by its capacity to evade phagocytosis, allowing dissemination in the host. Immune evasion requires a surface polysaccharide produced by all enterococci, known as the enterococcal polysaccharide antigen (EPA). EPA consists of a cell wall-anchored rhamnose backbone substituted by strain-specific polysaccharides called 'decorations', essential for the biological activity of this polymer. However, the structural determinants required for innate immune evasion remain unknown, partly due to a lack of suitable validated assays. Here, we describe a quantitative, in vitro assay to investigate how EPA decorations alter phagocytosis. Using the E. faecalis model strain OG1RF, we demonstrate that a mutant with a deletion of the locus encoding EPA decorations can be used as a platform strain to express heterologous decorations, thereby providing an experimental system to investigate the inhibition of phagocytosis by strain-specific decorations. We show that the aggregation of cells lacking decorations is increasing phagocytosis and that this process does not involve the recognition of lipoproteins by macrophages. Collectively, our work provides novel insights into innate immune evasion by enterococci and paves the way for further studies to explore the structure/function relationship of EPA decorations.
Assuntos
Enterococcus faecalis , Evasão da Resposta Imune , Lipoproteínas , Macrófagos , Fagocitose , Enterococcus faecalis/imunologia , Enterococcus faecalis/metabolismo , Enterococcus faecalis/genética , Lipoproteínas/metabolismo , Lipoproteínas/genética , Macrófagos/microbiologia , Macrófagos/imunologia , Macrófagos/metabolismo , Polissacarídeos Bacterianos/metabolismo , Polissacarídeos Bacterianos/imunologia , Humanos , Antígenos de Bactérias/metabolismo , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/genética , Imunidade Inata , Virulência , Animais , CamundongosRESUMO
The identification and recombinant production of functional antigens and/or epitopes of pathogens represent a crucial step for the development of an effective protein-based vaccine. Many vaccine targets are outer membrane proteins anchored into the lipidic bilayer through an extended hydrophobic portion making their recombinant production challenging. Moreover, only the extracellular loops, and not the hydrophobic regions, are naturally exposed to the immune system. In this work, the Domain 3 (D3) from Group B Streptococcus (GBS) pilus 2a backbone protein has been identified and engineered to be used as a scaffold for the display of extracellular loops of two Neisseria gonorrhoeae membrane proteins (PorB.1b and OpaB). A computational structure-based approach has been applied to the design of both the scaffold and the model antigens. Once identified the best D3 engineerable site, several different chimeric D3 displaying PorB.1b and OpaB extracellular loops were produced as soluble proteins. Each molecule has been characterized in terms of solubility, stability, and ability to correctly display the foreign epitope. This antigen dissection strategy allowed the identification of most immunogenic extracellular loops of both PorB.1b and OpaB gonococcal antigens. The crystal structure of chimeric D3 displaying PorB.1b immunodominant loop has been obtained confirming that the engineerization did not alter the predicted native structure of this epitope. Taken together, the reported data suggest that D3 is a novel protein scaffold for epitope insertion and display, and a valid alternative to the production of whole membrane protein antigens. Finally, this work describes a generalized computational structure-based approach for the identification, design, and dissection of epitopes in target antigens through chimeric proteins.
Assuntos
Proteínas de Membrana , Vacinas , Epitopos/genética , Antígenos de Bactérias/genética , Bicamadas LipídicasRESUMO
SignificanceYersinia pestis, the etiologic agent of plague, has been responsible for high mortality in several epidemics throughout human history. This plague bacillus has been used as a biological weapon during human history and is currently one of the deadliest biological threats. Currently, no licensed plague vaccines are available in the Western world. Since an array of immunogens are enclosed in outer membrane vesicles (OMVs), immune responses elicited by OMVs against a diverse range of antigens may reduce the likelihood of antigen circumvention. Therefore, self-adjuvanting OMVs from a remodeled Yersinia pseudotuberculosis strain as a type of plague vaccine could diversify prophylactic choices and solve current vaccine limitations.
Assuntos
Antígenos de Bactérias , Lipídeo A , Vacina contra a Peste , Peste , Proteínas Citotóxicas Formadoras de Poros , Yersinia pseudotuberculosis , Animais , Anticorpos Antibacterianos/sangue , Antígenos de Bactérias/genética , Antígenos de Bactérias/imunologia , Dose Letal Mediana , Lipídeo A/genética , Lipídeo A/imunologia , Camundongos , Peste/prevenção & controle , Vacina contra a Peste/administração & dosagem , Vacina contra a Peste/genética , Vacina contra a Peste/imunologia , Plasmídeos/genética , Proteínas Citotóxicas Formadoras de Poros/genética , Proteínas Citotóxicas Formadoras de Poros/imunologia , Yersinia pseudotuberculosis/genética , Yersinia pseudotuberculosis/imunologiaRESUMO
The bacterial genus Bartonella comprises numerous emerging pathogens that cause a broad spectrum of disease manifestations in humans. The targets and mechanisms of the anti-Bartonella immune defense are ill-defined and bacterial immune evasion strategies remain elusive. We found that experimentally infected mice resolved Bartonella infection by mounting antibody responses that neutralized the bacteria, preventing their attachment to erythrocytes and suppressing bacteremia independent of complement or Fc receptors. Bartonella-neutralizing antibody responses were rapidly induced and depended on CD40 signaling but not on affinity maturation. We cloned neutralizing monoclonal antibodies (mAbs) and by mass spectrometry identified the bacterial autotransporter CFA (CAMP-like factor autotransporter) as a neutralizing antibody target. Vaccination against CFA suppressed Bartonella bacteremia, validating CFA as a protective antigen. We mapped Bartonella-neutralizing mAb binding to a domain in CFA that we found is hypervariable in both human and mouse pathogenic strains, indicating mutational antibody evasion at the Bartonella subspecies level. These insights into Bartonella immunity and immune evasion provide a conceptual framework for vaccine development, identifying important challenges in this endeavor.
Assuntos
Anticorpos Neutralizantes , Antígenos de Bactérias , Bacteriemia , Infecções por Bartonella , Bartonella , Sistemas de Secreção Tipo V , Animais , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/genética , Anticorpos Neutralizantes/imunologia , Antígenos de Bactérias/genética , Antígenos de Bactérias/imunologia , Bacteriemia/imunologia , Bacteriemia/microbiologia , Bacteriemia/prevenção & controle , Vacinas Bacterianas/genética , Vacinas Bacterianas/imunologia , Vacinas Bacterianas/uso terapêutico , Bartonella/genética , Bartonella/imunologia , Infecções por Bartonella/imunologia , Infecções por Bartonella/microbiologia , Infecções por Bartonella/prevenção & controle , Clonagem Molecular , Evasão da Resposta Imune , Camundongos , Sistemas de Secreção Tipo V/imunologia , VacinaçãoRESUMO
SignificanceTuberculosis (TB), an ancient disease of humanity, continues to be a major cause of worldwide death. The causative agent of TB, Mycobacterium tuberculosis, and its close pathogenic relative Mycobacterium marinum, initially infect, evade, and exploit macrophages, a major host defense against invading pathogens. Within macrophages, mycobacteria reside within host membrane-bound compartments called phagosomes. Mycobacterium-induced damage of the phagosomal membranes is integral to pathogenesis, and this activity has been attributed to the specialized mycobacterial secretion system ESX-1, and particularly to ESAT-6, its major secreted protein. Here, we show that the integrity of the unstructured ESAT-6 C terminus is required for macrophage phagosomal damage, granuloma formation, and virulence.
Assuntos
Antígenos de Bactérias , Proteínas de Bactérias , Mycobacterium marinum , Mycobacterium tuberculosis , Fagossomos , Tuberculoma , Sistemas de Secreção Tipo VII , Antígenos de Bactérias/química , Antígenos de Bactérias/genética , Antígenos de Bactérias/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Humanos , Mycobacterium marinum/metabolismo , Mycobacterium marinum/patogenicidade , Mycobacterium tuberculosis/metabolismo , Mycobacterium tuberculosis/patogenicidade , Fagossomos/metabolismo , Fagossomos/microbiologia , Conformação Proteica , Tuberculoma/microbiologia , Sistemas de Secreção Tipo VII/metabolismo , VirulênciaRESUMO
Interactions among pathogen genotypes that vary in host specificity may affect overall transmission dynamics in multi-host systems. Borrelia burgdorferi, a bacterium that causes Lyme disease, is typically transmitted among wildlife by Ixodes ticks. Despite the existence of many alleles of B. burgdorferi's sensu stricto outer surface protein C (ospC) gene, most human infections are caused by a small number of ospC alleles ["human infectious alleles" (HIAs)], suggesting variation in host specificity associated with ospC. To characterize the wildlife host association of B. burgdorferi's ospC alleles, we used metagenomics to sequence ospC alleles from 68 infected individuals belonging to eight mammalian species trapped at three sites in suburban New Brunswick, New Jersey (USA). We found that multiple allele ("mixed") infections were common. HIAs were most common in mice (Peromyscus spp.) and only one HIA was detected at a site where mice were rarely captured. ospC allele U was exclusively found in chipmunks (Tamias striatus), and although a significant number of different alleles were observed in chipmunks, including HIAs, allele U never co-occurred with other alleles in mixed infections. Our results suggest that allele U may be excluding other alleles, thereby reducing the capacity of chipmunks to act as reservoirs for HIAs.
Assuntos
Borrelia burgdorferi , Borrelia , Coinfecção , Ixodes , Doença de Lyme , Animais , Humanos , Borrelia burgdorferi/genética , Borrelia/genética , Alelos , Doença de Lyme/microbiologia , Ixodes/genética , Ixodes/microbiologia , Antígenos de Bactérias/genética , Proteínas da Membrana Bacteriana Externa/genética , Sciuridae/genética , Especificidade de HospedeiroRESUMO
Extraintestinal pathogenic Escherichia coli (ExPEC) is a leading cause of worldwide morbidity and mortality, the top cause of antimicrobial-resistant (AMR) infections, and the most frequent cause of life-threatening sepsis and urinary tract infections (UTI) in adults. The development of an effective and universal vaccine is complicated by this pathogen's pan-genome, its ability to mix and match virulence factors and AMR genes via horizontal gene transfer, an inability to decipher commensal from pathogens, and its intimate association and co-evolution with mammals. Using a pan virulome analysis of >20,000 sequenced E. coli strains, we identified the secreted cytolysin α-hemolysin (HlyA) as a high priority target for vaccine exploration studies. We demonstrate that a catalytically inactive pure form of HlyA, expressed in an autologous host using its own secretion system, is highly immunogenic in a murine host, protects against several forms of ExPEC infection (including lethal bacteremia), and significantly lowers bacterial burdens in multiple organ systems. Interestingly, the combination of a previously reported autotransporter (SinH) with HlyA was notably effective, inducing near complete protection against lethal challenge, including commonly used infection strains ST73 (CFT073) and ST95 (UTI89), as well as a mixture of 10 of the most highly virulent sequence types and strains from our clinical collection. Both HlyA and HlyA-SinH combinations also afforded some protection against UTI89 colonization in a murine UTI model. These findings suggest recombinant, inactive hemolysin and/or its combination with SinH warrant investigation in the development of an E. coli vaccine against invasive disease.
Assuntos
Infecções por Escherichia coli , Proteínas de Escherichia coli , Vacinas contra Escherichia coli , Escherichia coli Extraintestinal Patogênica , Proteínas Hemolisinas , Animais , Escherichia coli Extraintestinal Patogênica/genética , Escherichia coli Extraintestinal Patogênica/imunologia , Infecções por Escherichia coli/prevenção & controle , Infecções por Escherichia coli/microbiologia , Infecções por Escherichia coli/imunologia , Camundongos , Proteínas Hemolisinas/imunologia , Proteínas Hemolisinas/genética , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/imunologia , Vacinas contra Escherichia coli/imunologia , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/genética , Feminino , Fatores de Virulência/genética , Fatores de Virulência/imunologia , Sistemas de Secreção Tipo V/imunologia , Sistemas de Secreção Tipo V/genética , Modelos Animais de Doenças , HumanosRESUMO
bb0616 of Borrelia burgdorferi, the Lyme disease pathogen, encodes a hypothetical protein of unknown function. In this study, we showed that BB0616 was not surface-exposed or associated with the membrane through localization analyses using proteinase K digestion and cell partitioning assays. The expression of bb0616 was influenced by a reduced pH but not by growth phases, elevated temperatures, or carbon sources during in vitro cultivation. A transcriptional start site for bb0616 was identified by using 5' rapid amplification of cDNA ends, which led to the identification of a functional promoter in the 5' regulatory region upstream of bb0616. By analyzing a bb0616-deficient mutant and its isogenic complemented counterparts, we found that the infectivity potential of the mutant was significantly attenuated. The inactivation of bb0616 displayed no effect on borrelial growth in the medium or resistance to oxidative stress, but the mutant was significantly more susceptible to osmotic stress. In addition, the production of global virulence regulators such as BosR and RpoS as well as virulence-associated outer surface lipoproteins OspC and DbpA was reduced in the mutant. These phenotypes were fully restored when gene mutation was complemented with a wild-type copy of bb0616. Based on these findings, we concluded that the hypothetical protein BB0616 is required for the optimal infectivity of B. burgdorferi, potentially by impacting B. burgdorferi virulence gene expression as well as survival of the spirochete under stressful conditions.
Assuntos
Proteínas de Bactérias , Borrelia burgdorferi , Regulação Bacteriana da Expressão Gênica , Doença de Lyme , Borrelia burgdorferi/genética , Borrelia burgdorferi/patogenicidade , Borrelia burgdorferi/metabolismo , Animais , Camundongos , Doença de Lyme/microbiologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Regiões Promotoras Genéticas , Fatores de Virulência/genética , Fatores de Virulência/metabolismo , Virulência , Camundongos Endogâmicos C3H , Fator sigma/genética , Fator sigma/metabolismo , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Sítio de Iniciação de Transcrição , Antígenos de Bactérias/genética , Antígenos de Bactérias/metabolismo , Teste de Complementação Genética , Concentração de Íons de HidrogênioRESUMO
The major gram-positive pathogen group A Streptococcus (GAS) is a model organism for studying microbial epidemics as it causes waves of infections. Since 1980, several GAS epidemics have been ascribed to the emergence of clones producing increased amounts of key virulence factors such as streptolysin O (SLO). Herein, we sought to identify mechanisms underlying our recently identified temporal clonal emergence among emm4 GAS, given that emergent strains did not produce augmented levels of virulence factors relative to historic isolates. By creating and analyzing isoallelic strains, we determined that a conserved mutation in a previously undescribed gene encoding a putative carbonic anhydrase was responsible for the defective in vitro growth observed in the emergent strains. We also identified that the emergent strains survived better inside macrophages and killed macrophages at lower rates than the historic strains. Via the creation of isogenic mutant strains, we linked the emergent strain "survival" phenotype to the downregulation of the SLO encoding gene and upregulation of the msrAB operon which encodes proteins involved in defense against extracellular oxidative stress. Our findings are in accord with recent surveillance studies which found a high ratio of mucosal (i.e., pharyngeal) relative to invasive infections among emm4 GAS. Since ever-increasing virulence is unlikely to be evolutionarily advantageous for a microbial pathogen, our data further understanding of the well-described oscillating patterns of virulent GAS infections by demonstrating mechanisms by which emergent strains adapt a "survival" strategy to outcompete previously circulating isolates.
Assuntos
Proteínas de Bactérias , Macrófagos , Infecções Estreptocócicas , Streptococcus pyogenes , Estreptolisinas , Fatores de Virulência , Streptococcus pyogenes/genética , Streptococcus pyogenes/patogenicidade , Streptococcus pyogenes/imunologia , Infecções Estreptocócicas/microbiologia , Infecções Estreptocócicas/imunologia , Infecções Estreptocócicas/mortalidade , Humanos , Macrófagos/microbiologia , Macrófagos/imunologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Estreptolisinas/genética , Estreptolisinas/metabolismo , Fatores de Virulência/genética , Mutação , Interações Hospedeiro-Patógeno/imunologia , Virulência/genética , Animais , Antígenos de Bactérias/genética , Antígenos de Bactérias/metabolismo , Antígenos de Bactérias/imunologia , Viabilidade Microbiana , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Camundongos , Regulação Bacteriana da Expressão Gênica , Proteínas de TransporteRESUMO
Pseudomonas aeruginosa poses a significant threat to immunocompromised individuals and those with cystic fibrosis. Treatment relies on antibiotics, but persistent infections occur due to intrinsic and acquired resistance of P. aeruginosa towards multiple classes of antibiotics. To date, there are no licensed vaccines for this pathogen, prompting the urgent need for novel treatment approaches to combat P. aeruginosa infection and persistence. Here we validated AAV vectored immunoprophylaxis as a strategy to generate long-term plasma and mucosal expression of highly protective monoclonal antibodies (mAbs) targeting the exopolysaccharide Psl (Cam-003) and the PcrV (V2L2MD) component of the type-III secretion system injectosome either as single mAbs or together as a bispecific mAb (MEDI3902) in a mouse model. When administered intramuscularly, AAV-αPcrV, AAV-αPsl, and AAV-MEDI3902 significantly protected mice challenged intranasally with a lethal dose of P. aeruginosa strains PAO1 and PA14 and reduced bacterial burden and dissemination to other organs. While all AAV-mAbs provided protection, AAV-αPcrV and AAV-MEDI3902 provided 100% and 87.5% protection from a lethal challenge with 4.47 × 107 CFU PAO1 and 87.5% and 75% protection from a lethal challenge with 3 × 107 CFU PA14, respectively. Serum concentrations of MEDI3902 were ~10× lower than that of αPcrV, but mice treated with this vector showed a greater reduction in bacterial dissemination to the liver, lung, spleen, and blood compared to other AAV-mAbs. These results support further investigation into the use of AAV vectored immunoprophylaxis to prevent and treat P. aeruginosa infections and other bacterial pathogens of public health concern for which current treatment strategies are limited.
Assuntos
Anticorpos Monoclonais , Dependovirus , Vetores Genéticos , Infecções por Pseudomonas , Pseudomonas aeruginosa , Animais , Dependovirus/genética , Dependovirus/imunologia , Pseudomonas aeruginosa/imunologia , Pseudomonas aeruginosa/genética , Camundongos , Vetores Genéticos/genética , Vetores Genéticos/administração & dosagem , Infecções por Pseudomonas/prevenção & controle , Infecções por Pseudomonas/imunologia , Infecções por Pseudomonas/terapia , Anticorpos Monoclonais/imunologia , Anticorpos Biespecíficos , Feminino , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/genética , Pneumonia Bacteriana/prevenção & controle , Pneumonia Bacteriana/terapia , Pneumonia Bacteriana/imunologia , Modelos Animais de Doenças , Camundongos Endogâmicos C57BL , Humanos , Toxinas Bacterianas , Proteínas Citotóxicas Formadoras de PorosRESUMO
As a potential side effect of the severe acute respiratory syndrome coronavirus type 2 pandemic, invasive group A Streptococcus (iGAS) infections in Europe have increased dramatically in both children and adults in the end of 2022. This epidemiological and molecular study describes the distributions of streptococcal genes encoding the M antigen (emm types) and superantigens in patients with invasive and non-invasive GAS infections. From December 2022 to December 2023, a total of 163 GAS isolates were collected from sterile and non-sterile sites of patients at five hospitals in Germany including two tertiary care centers. Genes encoding M protein and superantigens were determined following the guidelines of CDC Streptococcus laboratory. Patients' characteristics were reviewed retrospectively. Correlations of clinical factors, emm types, and superantigens with rates of invasive infections were analyzed. Of the 163 included GAS cases, 112 (69%) were considered as invasive. In total, 33 different emm types were observed, of which emm1.0 (n = 49; 30%), emm89.0 (n = 15; 9%), and emm12.0 (n = 14; 9%) were most prevalent. In total, 70% of emm1.0 isolates belonged to M1UK lineage. No difference in invasive infections was observed for the M1UK lineage compared with other emm1.0 isolates. However, the emm1.0 type, presence of speA1-3, speG, or speJ, as well as adulthood were significantly associated with invasive infections. In contrast, emm12.0 isolates were significantly less associated with invasive infections. Multivariable analysis confirmed a significant influence of speJ and adulthood on iGAS infections. This study underlines the importance of continuous monitoring of genomic trends and identification of emerging GAS variants. This may aid in delineating pathogenicity factors of Streptococcus pyogenes that propel invasive infections.
Assuntos
Antígenos de Bactérias , Proteínas da Membrana Bacteriana Externa , Proteínas de Transporte , Infecções Estreptocócicas , Streptococcus pyogenes , Humanos , Infecções Estreptocócicas/microbiologia , Infecções Estreptocócicas/epidemiologia , Streptococcus pyogenes/genética , Streptococcus pyogenes/classificação , Streptococcus pyogenes/isolamento & purificação , Alemanha/epidemiologia , Estudos Retrospectivos , Proteínas da Membrana Bacteriana Externa/genética , Adulto , Feminino , Masculino , Pessoa de Meia-Idade , Criança , Antígenos de Bactérias/genética , Proteínas de Transporte/genética , Adolescente , Pré-Escolar , Idoso , Adulto Jovem , Lactente , Superantígenos/genética , Idoso de 80 Anos ou maisRESUMO
Helicobacter pylori (H. pylori) is a common gastric pathogen that infects approximately half of the world's population. Infection with H. pylori can lead to diverse pathological conditions, including chronic gastritis, peptic ulcer disease, and cancer. The latter is the most severe consequence of H. pylori infection. According to epidemiological studies, gastric infection with H. pylori is the strongest known risk factor for non-cardia gastric cancer (GC), which remains one of the leading causes of cancer-related deaths worldwide. However, it still remains to be poorly understood how host-microbe interactions result in cancer development in the human stomach. Here we focus on the H. pylori bacterial factors that affect the host ubiquitin proteasome system. We investigated E3 ubiquitin ligases SIVA1 and ULF that regulate p14ARF (p19ARF in mice) tumor suppressor. ARF plays a key role in regulation of the oncogenic stress response and is frequently inhibited during GC progression. Expression of ARF, SIVA1 and ULF proteins were investigated in gastroids, H. pylori-infected mice and human gastric tissues. The role of the H. pylori type IV secretion system was assessed using various H. pylori isogenic mutants. Our studies demonstrated that H. pylori infection results in induction of ULF, decrease in SIVA1 protein levels, and subsequent ubiquitination and degradation of p14ARF tumor suppressor. Bacterial CagA protein was found to sequentially bind to SIVA1 and ULF proteins. This process is regulated by CagA protein phosphorylation at the EPIYA motifs. Downregulation of ARF protein leads to inhibition of cellular apoptosis and oncogenic stress response that may promote gastric carcinogenesis.
Assuntos
Infecções por Helicobacter , Helicobacter pylori , Neoplasias Gástricas , Animais , Antígenos de Bactérias/genética , Antígenos de Bactérias/metabolismo , Apoptose , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Carcinogênese/metabolismo , Mucosa Gástrica/microbiologia , Infecções por Helicobacter/microbiologia , Helicobacter pylori/metabolismo , Camundongos , Neoplasias Gástricas/metabolismo , Proteína Supressora de Tumor p14ARF/metabolismo , Ubiquitinas/metabolismoRESUMO
Hematogenous dissemination is a critical step in the evolution of local infection to systemic disease. The Lyme disease (LD) spirochete, which efficiently disseminates to multiple tissues, has provided a model for this process, in particular for the key early event of pathogen adhesion to the host vasculature. This occurs under shear force mediated by interactions between bacterial adhesins and mammalian cell-surface proteins or extracellular matrix (ECM). Using real-time intravital imaging of the Lyme spirochete in living mice, we previously identified BBK32 as the first LD spirochetal adhesin demonstrated to mediate early vascular adhesion in a living mouse; however, deletion of bbk32 resulted in loss of only about half of the early interactions, suggesting the existence of at least one other adhesin (adhesin-X) that promotes early vascular interactions. VlsE, a surface lipoprotein, was identified long ago by its capacity to undergo rapid antigenic variation, is upregulated in the mammalian host and required for persistent infection in immunocompetent mice. In immunodeficient mice, VlsE shares functional overlap with OspC, a multi-functional protein that displays dermatan sulfate-binding activity and is required for joint invasion and colonization. In this research, using biochemical and genetic approaches as well as intravital imaging, we have identified VlsE as adhesin-X; it is a dermatan sulfate (DS) adhesin that efficiently promotes transient adhesion to the microvasculature under shear force via its DS binding pocket. Intravenous inoculation of mice with a low-passage infectious B. burgdorferi strain lacking both bbk32 and vlsE almost completely eliminated transient microvascular interactions. Comparative analysis of binding parameters of VlsE, BBK32 and OspC provides a possible explanation why these three DS adhesins display different functionality in terms of their ability to promote early microvascular interactions.
Assuntos
Adesinas Bacterianas , Variação Antigênica , Antígenos de Bactérias , Proteínas de Bactérias , Borrelia burgdorferi , Lipoproteínas , Doença de Lyme , Microvasos , Adesinas Bacterianas/genética , Adesinas Bacterianas/imunologia , Animais , Variação Antigênica/genética , Variação Antigênica/imunologia , Antígenos de Bactérias/genética , Antígenos de Bactérias/imunologia , Aderência Bacteriana/genética , Aderência Bacteriana/imunologia , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/imunologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Borrelia burgdorferi/genética , Borrelia burgdorferi/imunologia , Dermatan Sulfato/imunologia , Lipoproteínas/genética , Lipoproteínas/imunologia , Doença de Lyme/genética , Doença de Lyme/imunologia , Doença de Lyme/microbiologia , Mamíferos , Camundongos , Microvasos/imunologia , Microvasos/microbiologia , Resistência ao CisalhamentoRESUMO
BACKGROUND: Enterococcus faecium and Staphylococcus aureus are the Gram-positive pathogens of the ESKAPE group, known to represent a great threat to human health due to their high virulence and multiple resistances to antibiotics. Combined, enterococci and S. aureus account for 26% of healthcare-associated infections and are the most common organisms responsible for blood stream infections. We previously showed that the peptidyl-prolyl cis/trans isomerase (PPIase) PpiC of E. faecium elicits the production of specific, opsonic, and protective antibodies that are effective against several strains of E. faecium and E. faecalis. Due to the ubiquitous characteristics of PPIases and their essential function within Gram-positive cells, we hypothesized a potential cross-reactive effect of anti-PpiC antibodies. RESULTS: Opsonophagocytic assays combined with bioinformatics led to the identification of the foldase protein PrsA as a new potential vaccine antigen in S. aureus. We show that PrsA is a stable dimeric protein able to elicit opsonic antibodies against the S. aureus strain MW2, as well as cross-binding and cross-opsonic in several S. aureus, E. faecium and E. faecalis strains. CONCLUSIONS: Given the multiple antibiotic resistances S. aureus and enterococci present, finding preventive strategies is essential to fight those two nosocomial pathogens. The study shows the potential of PrsA as an antigen to use in vaccine formulation against the two dangerous Gram-positive ESKAPE bacteria. Our findings support the idea that PPIases should be further investigated as vaccine targets in the frame of pan-vaccinomics strategy.
Assuntos
Proteínas de Bactérias , Enterococcus faecalis , Enterococcus faecium , Peptidilprolil Isomerase , Staphylococcus aureus , Staphylococcus aureus/imunologia , Staphylococcus aureus/genética , Enterococcus faecium/imunologia , Enterococcus faecium/genética , Proteínas de Bactérias/imunologia , Proteínas de Bactérias/genética , Peptidilprolil Isomerase/imunologia , Peptidilprolil Isomerase/genética , Enterococcus faecalis/imunologia , Enterococcus faecalis/genética , Humanos , Infecções por Bactérias Gram-Positivas/prevenção & controle , Infecções por Bactérias Gram-Positivas/imunologia , Infecções por Bactérias Gram-Positivas/microbiologia , Vacinas Bacterianas/imunologia , Proteínas Opsonizantes/imunologia , Anticorpos Antibacterianos/imunologia , Anticorpos Antibacterianos/sangue , Animais , Reações Cruzadas , Camundongos , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/genética , Fagocitose , Infecções Estafilocócicas/prevenção & controle , Infecções Estafilocócicas/imunologia , Infecções Estafilocócicas/microbiologiaRESUMO
The PE and PPE family proteins of Mycobacterium tuberculosis (Mtb) is exclusively found in pathogenic Mycobacterium species, comprising approximately 8-10 % of the Mtb genome. These emerging virulent factors have been observed to play pivotal roles in Mtb pathogenesis and immune evasion through various strategies. These immunogenic proteins are known to modulate the host immune response and cell-death pathways by targeting the powerhouse of the cell, the mitochondria to support Mtb survival. In this article, we are focused on how PE/PPE family proteins target host mitochondria to induce mitochondrial perturbations, modulate the levels of cellular ROS (Reactive oxygen species) and control cell death pathways. We observed that the time of expression of these proteins at different stages of infection is crucial for elucidating their impact on the cell death pathways and eventually on the outcome of infection. This article focuses on understanding the contributions of the PE/PPE proteins by unravelling the triad of host mitochondria, oxidative stress and cell death pathways that facilitate the Mtb persistence. Understanding the role of these proteins in host cellular pathways and the intricate mechanisms paves the way for the development of novel therapeutic strategies to combat TB infections.
Assuntos
Proteínas de Bactérias , Morte Celular , Interações Hospedeiro-Patógeno , Mitocôndrias , Mycobacterium tuberculosis , Espécies Reativas de Oxigênio , Humanos , Antígenos de Bactérias/metabolismo , Antígenos de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Mitocôndrias/metabolismo , Mycobacterium tuberculosis/patogenicidade , Mycobacterium tuberculosis/metabolismo , Mycobacterium tuberculosis/genética , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Tuberculose/microbiologia , Tuberculose/metabolismo , Fatores de Virulência/metabolismoRESUMO
Enterotoxigenic Escherichia coli (ETEC) is one of the main causes of diarrhea in children and travelers in low-income regions. The virulence of ETEC is attributed to its heat-labile and heat-stable enterotoxins, as well as its colonization factors (CFs). CFs are essential for ETEC adherence to the intestinal epithelium. However, its invasive capability remains unelucidated. In this study, we demonstrated that the CS6-positive ETEC strain 4266 can invade mammalian epithelial cells. The invasive capability was reduced in the 4266 ΔCS6 mutant but reintroduction of CS6 into this mutant restored the invasiveness. Additionally, the laboratory E. coli strain Top 10, which lacks the invasive capability, was able to invade Caco-2 cells after gaining the CS6-expressing plasmid pCS6. Cytochalasin D inhibited cell invasion in both 4266 and Top10 pCS6 cells, and F-actin accumulation was observed near the bacteria on the cell membrane, indicating that CS6-positive bacteria were internalized via actin polymerization. Other cell signal transduction inhibitors, such as genistein, wortmannin, LY294002, PP1, and Ro 32-0432, inhibited the CS6-mediated invasion of Caco-2 cells. The internalized bacteria of both 4266 and Top10 pCS6 strains were able to survive for up to 48 h, and 4266 cells were able to replicate within Caco-2 cells. Immunofluorescence microscopy revealed that the internalized 4266 cells were present in bacteria-containing vacuoles, which underwent a maturation process indicated by the recruitment of the early endosomal marker EEA-1 and late endosomal marker LAMP-1 throughout the infection process. The autophagy marker LC3 was also observed near these vacuoles, indicating the initiation of LC-3-associated phagocytosis (LAP). However, intracellular bacteria continued to replicate, even after the initiation of LAP. Moreover, intracellular filamentation was observed in 4266 cells at 24 h after infection. Overall, this study shows that CS6, in addition to being a major CF, mediates cell invasion. This demonstrates that once internalized, CS6-positive ETEC is capable of surviving and replicating within host cells. This capability may be a key factor in the extended and recurrent nature of ETEC infections in humans, thus highlighting the critical role of CS6.
Assuntos
Citocalasina D , Escherichia coli Enterotoxigênica , Proteínas de Escherichia coli , Humanos , Células CACO-2 , Escherichia coli Enterotoxigênica/patogenicidade , Escherichia coli Enterotoxigênica/genética , Escherichia coli Enterotoxigênica/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Citocalasina D/farmacologia , Actinas/metabolismo , Células Epiteliais/microbiologia , Aderência Bacteriana , Infecções por Escherichia coli/microbiologia , Virulência , Fatores de Virulência/genética , Fatores de Virulência/metabolismo , Antígenos de Bactérias/metabolismo , Antígenos de Bactérias/genética , Morfolinas/farmacologia , Transdução de Sinais , Androstadienos/farmacologia , Wortmanina/farmacologia , Endocitose , Cromonas/farmacologia , Plasmídeos/genéticaRESUMO
QseC is a membrane sensor kinase that enables bacteria to perceive autoinducers -3, adrenaline, and norepinephrine to initiate downstream gene transcription. In this study, we found that the QseC protein of Glaesserella parasuis can serve as an effective antigen to activate the host's immune response. Therefore, we investigated the immunogenicity and host protective effect of this protein. ELISA and indirect immunofluorescence results showed that QseC protein can induce high titer levels of humoral immunity in mice and regularly generate specific serum antibodies. We used MTS reagents to detect lymphocyte proliferation levels and found that QseC protein can cause splenic lymphocyte proliferation with memory and specificity. Further immunological analysis of the spleen cell supernatant revealed significant upregulation of levels of IL-1ß, IL-4 and IFN-γ in the QseC + adjuvant group. In the mouse challenge experiment, it was found that QseC + adjuvant can provide effective protection. The results of this study demonstrate that QseC protein provides effective protection in a mouse model and has the potential to serve as a candidate antigen for a novel subunit vaccine for further research.
Assuntos
Anticorpos Antibacterianos , Infecções por Haemophilus , Interferon gama , Interleucina-4 , Animais , Camundongos , Interleucina-4/metabolismo , Interleucina-4/imunologia , Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/imunologia , Infecções por Haemophilus/imunologia , Infecções por Haemophilus/prevenção & controle , Infecções por Haemophilus/microbiologia , Interferon gama/metabolismo , Histidina Quinase/genética , Histidina Quinase/metabolismo , Histidina Quinase/imunologia , Interleucina-1beta/metabolismo , Interleucina-1beta/genética , Imunidade Humoral , Camundongos Endogâmicos BALB C , Baço/imunologia , Proteínas de Bactérias/imunologia , Proteínas de Bactérias/genética , Proliferação de Células , Feminino , Adjuvantes Imunológicos , Haemophilus parasuis/imunologia , Haemophilus parasuis/genética , Citocinas/metabolismo , Vacinas Bacterianas/imunologia , Vacinas Bacterianas/genética , Modelos Animais de Doenças , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/genética , Linfócitos/imunologia , Vacinas de Subunidades Antigênicas/imunologia , Vacinas de Subunidades Antigênicas/genéticaRESUMO
Streptococcus suis is one of the major pathogens of pigs circulating worldwide, and the development of vaccines will help to effectively control streptococcosis in swine. In this study, we evaluated the potential of three membrane associated proteins, histidine kinase (HK), glycosyltransferase family 2 (Gtf-2) and phosphate binding protein (PsbP) of S. suis as subunit vaccines. Bioinformatics analysis shows that protein ABC is highly conserved in S. suis. To verify the protective effects of these proteins in animal models, recombinant protein HK, Gtf-2 and PsbP were used to immunize BALB/c mice separately. The results showed that these proteins immunization in mice can effectively induce strong humoral immune responses, protect mice from cytokine storms caused by S. suis infection, and have a significant protective effect against lethal doses of S. suis infection. Furthermore, antibodies with opsonic activity exist in the recombinant proteins antiserum to assist phagocytic cells in killing S. suis. Overall, these results indicated that these recombinant proteins all elicit good immune protective effect against S. suis infection and can be represent promising candidate antigens for subunit vaccines against S. suis.